Project Summary Targeting the androgen receptor (AR) is the mainstay of treatment for metastatic prostate cancer. This usually involves either LHRH agonists that prevent testicular androgen synthesis or AR antagonists, such as bicalutamide (Casodex), which block AR transcriptional activity. Although initial responses to these therapies are effective, they inevitably fail because castration-resistant prostate cancer cells emerge with enhanced AR activity. Thus, castration-resistant prostate cancers maintain their reliance on androgen signaling. This prompted the development of a new generation of anti-androgens, such as enzalutamide, which blocks AR action by inhibiting translocation of AR into the nucleus. Although enzalutamide represents a breakthrough in treatment of metastatic prostate cancer, patients who initially respond eventually acquire resistance. Given that one mechanism of enzalutamide resistance relies on the ability of the glucocorticoid receptor (GR) to substitute for AR to activate a similar set of target genes necessary for proliferation, blocking GR activity might combat certain cases of enzalutamide resistance. We have previously identified a set of 23 cellular factors that reduce GR-dependent transcriptional activation when depleted. We hypothesize that these factors dictate GR specificity at gene targets in enzalutamide-resistant prostate cancer cells. We further propose that targeting these factors would block the proliferation of GR-driven, enzalutamide-resistant prostate cancer. Our specific aims are 1) to define the impact of these GR cofactors on GR-expressing, enzalutamide-resistant prostate cancer cells and 2) to determine the capacity of the GR cofactors to inhibit enzalutamide-resistant tumor xenograft growth in vivo. Successful completion of these aims will identify key regulators of GR transcription and proliferation in enzalutamide-resistant prostate cancer.